Endothelial cell substrate stiffness influences neutrophil transmigration via myosin light chain kinase-dependent cell contraction

Blood. 2011 Aug 11;118(6):1632-40. doi: 10.1182/blood-2010-11-321125. Epub 2011 Jun 7.

Abstract

A vast amount of work has been dedicated to the effects of shear flow and cytokines on leukocyte transmigration. However, no studies have explored the effects of substrate stiffness on transmigration. Here, we investigated important aspects of endothelial cell contraction-mediated neutrophil transmigration using an in vitro model of the vascular endothelium. We modeled blood vessels of varying mechanical properties using fibronectin-coated polyacrylamide gels of varying physiologic stiffness, plated with human umbilical vein endothelial cell (HUVEC) monolayers, which were activated with tumor necrosis factor-α. Interestingly, neutrophil transmigration increased with increasing substrate stiffness below the endothelium. HUVEC intercellular adhesion molecule-1 expression, stiffness, cytoskeletal arrangement, morphology, and cell-substrate adhesion could not account for the dependence of transmigration on HUVEC substrate stiffness. We also explored the role of cell contraction and observed that large holes formed in endothelium on stiff substrates several minutes after neutrophil transmigration reached a maximum. Further, suppression of contraction through inhibition of myosin light chain kinase normalized the effects of substrate stiffness by reducing transmigration and eliminating hole formation in HUVECs on stiff substrates. These results provide strong evidence that neutrophil transmigration is regulated by myosin light chain kinase-mediated endothelial cell contraction and that this event depends on subendothelial cell matrix stiffness.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acrylic Resins / metabolism
  • Azepines / pharmacology
  • Cell Adhesion / physiology
  • Cell Shape / physiology
  • Cells, Cultured
  • Endothelial Cells / drug effects
  • Endothelial Cells / metabolism*
  • Enzyme Inhibitors / pharmacology
  • Fibronectins / metabolism
  • Humans
  • Immunohistochemistry
  • Intercellular Adhesion Molecule-1 / metabolism
  • Microscopy, Atomic Force
  • Myosin-Light-Chain Kinase / antagonists & inhibitors
  • Myosin-Light-Chain Kinase / metabolism*
  • Naphthalenes / pharmacology
  • Neutrophils / physiology*
  • Transendothelial and Transepithelial Migration / drug effects
  • Transendothelial and Transepithelial Migration / physiology*
  • Tumor Necrosis Factor-alpha / pharmacology

Substances

  • Acrylic Resins
  • Azepines
  • Enzyme Inhibitors
  • Fibronectins
  • Naphthalenes
  • Tumor Necrosis Factor-alpha
  • polyacrylamide gels
  • ML 7
  • Intercellular Adhesion Molecule-1
  • Myosin-Light-Chain Kinase